Investigators have shown an intense interest in organic conducting polymers that can be synthesized chemically, like polyacetylene, or electrochemically, like polypyrrole and polythiophene. The organic conducting polymers have several potential applications in the fields of batteries, display devices, corrosion prevention in metals and semiconductors and in microelectronic devices such as diodes, transistors, sensors, light emitting devices and energy conversion and storage elements. However, present day organic conducting polymers possess several limitations that have hindered the expansion of organic conducting polymers into these and other potential application areas. The limitations found in the three most extensively studied conducting polymers, polyacetylene, polypyrrole and polythiophene, illustrate the general problems encountered by investigators in the field of conducting polymers and why the use of conducting polymers has been impeded.
For example, polyacetylene, among the first organic conducting polymers, is prepared chemically from acetylene by using an appropriate catalyst. As prepared chemically, polyacetylene is an insulator, exhibiting conductivities in the range of 10.sup.-10 S/cm to 10.sup.-13 S/cm (Siemens per centimeter) that correspond to the conductivity of known insulators, such as glass and DNA. However, polyacetylene can be doped using a variety of oxidizing or reducing agents, such as antimony pentafluoride, the halogens, astatine pentafluoride, or aluminum chloride. By doping, polyacetylene is converted into a highly conducting polymer, exhibiting a conductivity of approximately 10.sup.3 S/cm, therefore exhibiting the conductivity of metals such as bismuth. However, polyacetylene suffers from the drawbacks of extreme instability in air and a precipitous drop in conductivity whenever an acetylenic hydrogen is replaced by an alkyl or other substituent group. Accordingly, the instability of polyacetylene in the presence of oxygen, and its inability to be functionalized and maintain its high conductivity, makes the polyacetylenes unsuitable conducting polymers for use as an analyte sensor.
Polypyrrole, a conducting polymer similar to polyacetylene, can be synthesized chemically or electrochemically and exhibits conductivities ranging form about 1 S/cm to about 100 S/cm. As will be discussed more fully hereinafter, conducting polypyrrole is a doped material, incorporating the anion of the supporting electrolyte. Polypyrrole having a molecular weight of up to approximately 40,000 has been synthesized; however, conductivity is observed in polypyrrole containing as few as six monomer units. Normally, polypyrrole, and other conducting polymers, are low molecular weight polymers containing less than 100 monomer units.
Investigators have found that placing alkyl groups on either the nitrogen or the carbons of the heteroaromatic pyrrole ring decreases the conductivity of polypyrrole. For example, an unsubstituted polypyrrole, incorporating the tetrafluoroborate anion as the compensating counterion, exhibits a conductivity of 40 S/cm, whereas the N-methyl derivative, incorporating the same dopant, exhibits a conductivity of 10.sup.-3 S/cm; the three-methyl derivative of pyrrole exhibits a conductivity of 4 S/cm; 3,4-dimethyl derivative, a conductive of 10 S/cm; and the 3,4-diphenyl derivative, a conductivity of 10.sup.-3 S/cm.
The conductivity decrease in substituted polypyrroles is attributed to several factors. First, and of prime importance, the substituent introduced onto the heteroaromatic pyrrole ring cannot alter the oxidation potential of the parent heteroaromatic to the extent that electropolymerization at the anode is precluded. Secondly, and a related consideration, the aromatic pi-electron system of the parent heterocycle must be maintained. Disruption of the pi-electron system of the heteroaromatic ring will adversely affect the relative stability of the aromatic and quinoid-like forms, illustrated as structures I and II, respectively, and therefore seriously reduce conductivity. A third critical consideration is that the functionality introduced onto the parent heterocycle must not create steric demands that preclude the adoption of a planar configuration by the conducting polymer. ##STR1##
The requirement that the conducting polymer must maintain a planar configuration has seriously hindered development of functionalized, conducting polymers. Numerous N-alkyl and N-aryl derivatives of polypyrrole have been prepared and discussed in the literature. However, it was found that even the simplest of these N-substituted polypyrroles, poly-N-methylpyrrole, exhibits conductivities that are three orders of magnitude lower than unsubstituted polypyrrole films doped with the same counterion. It is also possible to produce thin films of poly-N-aryl pyrroles, wherein the phenyl group is further substituted in the para position. However, polymers produced from these N-aryl pyrroles invariably exhibit conductivities three or more orders of magnitude less than the parent unsubstituted pyrrole. Such low conductivities preclude the use of these substituted polypyrroles in the development of analyte sensors.
The steric interactions introduced by the pyrrole ring substituents is important because of the mechanism of charge transport through the conducting polymer system. In one charge transport mechanism, electric charge is conducted through the polymer chain itself because of bipolaron structures that exist along the polymer chain. The bipolaron structure, illustrated in structure III and confirmed from spectroscopic evidence obtained on polythiophene, are defects occurring in the polymer lattice wherein two dopant counterions, A.sup.-, from the supporting electrolyte, balance two positive centers found in the polymer. ##STR2##
Generally, the two positive centers are spaced, and confined, by approximately four monomer units and these defects serve to transport charge along the polymer chain. However, in order to transport charge along the chain, compositions having the structures I, II and/or III must be planar, such that the charge can be transported along the planar pi-electron system of the chain. As can be seen in structure IV, if the substituents R and/or R' are sufficiently large, the steric interaction between R and R' can distort the pyrrole monomer units out of planarity, therefore destroying the planarity of the pi-electron system, and destroying, or seriously reducing, the conductivity of the polymer. As illustrated by the large conductivity drop in polypyrroles having substituents positioned on the pyrrole ring, even substituents as small as a methyl group introduce steric interactions sufficient to essentially destroy the conductivity of the polymer.
It also should be noted that investigators have found that R and/or R' substituents in structure IV should not be strongly electron-withdrawing or strongly electron-donating, as strong electronic effects also can serve to destroy the conductivity of the polymer. However, it has been found, especially for N-substituted pyrroles, that steric interactions, not electronic effects, are the main factor in determining polymerizability, polymer conductivity, and cyclic stability of the polymer between the doped and undoped state. Steric interactions in polythiophene derivatives are somewhat less dominant than those observed in polypyrrole derivatives. Steric interactions in polypyrrole derivatives are more dominant because the predominant destabilizing interactions in pyrrole derivatives involve the hydrogen atom of the pyrrole nitrogen. These steric interactions are avoided in polythiophenes. As a result, electronic effects play a more central role in polythiophene derivatives.
Conducting organic polymers generally are amorphous, disordered materials, and as a result, if bulk conductivity is to be sustained, charge transport must occur between polymer strands as well as along single polymer strands. The probability of the interchain charge transport is directly related to the distance between chains. The distance between polymer chains is acutely sensitive to, and dependent upon, two factors, the nature and size of the dopant counterion and the character and steric requirements of the R and R' substituents of structure IV. The steric requirement imposes a significant constraint on the design of functionalized conducting polymers.
The synthesis and conductivities of polypyrrole and substituted polypyrroles have been extensively investigated as seen in the general reference cited below. These references include the information discussed above and general information concerning the polypyrroles, such as that the specific dopant (A.sup.-) in structure III can seriously affect the conductivity of the polymer; that conductivity is observed only for alpha-alpha coupling of monomers and not for alpha-beta coupling of monomers (see structure V); and that polypyrrole films are stable, insoluble, and inert to most reagents, except possibly treatment by alkalis. The conductivity and stability of polypyrrole makes polypyrrole a good candidate for use in analyte sensors, if the polypyrrole conductivity can be maintained when functional groups are introduced onto the heteroaromatic ring. ##STR3##
The representative references discussing the polypyrroles include:
G. Bidan, Tet. Lett. 26(6), 735-6 (1975). PA0 P. Audebert, G. Bidan, an M. Lapowski, J.C.S. Chem. Comm., 887 (1986). PA0 M. S. Wrighton, Science 231, 32 (1986). PA0 R. A. Simon, A. I. Ricco and M. S. Wrighton, J. Am. Chem. Soc, 104, 2034 (1982). PA0 A. F. Diaz, J. Castillo, K. K. Kanazawa, J. A. Logan, M. Salmon and O. Fojards, J. Electroanal. Chem. 133, 233 (1982). PA0 M. Saloma, M. Aguilar and M. Salmon, J. Electrochem. Soc. 132, 2379 (1985). PA0 M. V. Rosenthal, T. A. Skotheim, A. Melo, M. I. Florit, and M. Salmon, J. Electroanal. Chem. and Interfac. Chem. 1, 297 (1985). PA0 G. Bidan and M. Guglielmi, Synth. Met. 15, 51 (1986). PA0 M. Salmon and G. Bidan, J. Electrochem. Soc., 1897 (1985). PA0 E. M. Genies and A. A. Syed, Synth. Met. 10, 27 (1984/85). PA0 G. Bidan, A. Deronzier and J. C. Moutet, Nouveau Jour. de Chimie 8, 501 (1984). PA0 J. P. Travers, P. Audebert and G. Bidan, Mol. Cryst. Liq. Cryst. 118, 149 (1985). PA0 G. Tourillon, "Handbook of Conducting Polymers," T. A. Skotheim, ed., Marcel Dekker, Inc., New York, 1986, p. 193. PA0 R. J. Waltham, J. Bargon and A. F. Diaz, J. Phys. Chem. 87, 1459 (1983). PA0 G. Tourillon and F. Garnier, J. Polym. Sci. Polym. Phys. Ed. 22, 33 (1984). PA0 G. Tourillon and F. Garnier, J. Electroanal. Chem. 161, 51 (1984). PA0 A. F. Diaz and J. Bargon, "Handbook of Conducting Polymers," T. A. Skotheim, ed., Marcel Dekker, Inc., New York 1986, p. 81. PA0 J. Bargon, S. Mohmand and R. J. Waltman, IBM, J. Res. Dev. 27, 330 (1983). PA0 G. Tourillon and F. Garnier, J. Phys. Chem. 87, 2289 (1983). PA0 A. Czerwinski, H. Zimmer, C. H. Pham, and H. B. Mark, Jr., J. Electrochem. Soc. 132, 2669 (1985). PA0 Y. Ikariyama and W. R. Heineman, Anal. Chem. 58, 1803 (1986). PA0 M. Josowicz and J. Janata, Anal. Chem. 58, 514 (1986). PA0 T. N. Misra, B. Rosenberg and R. Switzer, J. Chem. Phys. 48, 2096 (1968). PA0 K. Yoshino, H. S. Nalwa, J. G. Rabe and W. F. Schmidt, Polymer Comm. 26, 103 (1985). PA0 C. Nylander, M. Armgrath and I. Lundstrom, Anal. Chem. Symp. Ser. 17 (Chem Sens) 159 (1983). PA0 H. S. White, G. P. Kittlesen and M. S. Wrighton, J. Am. Chem. Soc. 106, 5317 (1984). PA0 G. P. Kittlesen, H. S. White and M. S. Wrighton, J. Am. Chem. Soc. 106, 7389 (1984). PA0 A. S. Davydov, J. Theor. Biol. 38, 559 (1973). PA0 A. S. Davydov, Physica. Scripta. 20, 387 (1979). PA0 A. S. Davydov, studia biophysica (Berlin) 62, 1 (1977). PA0 A. C. Scott, "Nonlinear Electrodynamics in Biological Systems," M. Ross Adey and A. L. Lawrence, eds., Plenum Press, NY, 1984, p. 133. PA0 C. F. McClare, Nature 296, 88 (1972). PA0 H. Weinberg and J. Metselur, Syn, Comm. 14(1), 1 (1984). PA0 (1) R. Huisgen, H. Gotthardt and H. O. Bayer, Chem. Ber. 103, 2368 (1970). PA0 (2) J. W. Lown and B. E. Landberg, Can. J. Chem. 52, 798 (1974).
Another well-studied conducting polymer is polythiophene, wherein thiophene (structure V, X=S) is electrochemically polymerized to yield a stable conducting polymer. Similarly, furan (structure V, X=O) also yields a stable conducting polymer similar to polypyrrole and polythiophene. Polythiophene resembles polypyrrole in that polythiophene can be cyclized between its conducting (oxidized) state and its nonconducting (neutral) state without significant chemical decomposition of the polymer and without appreciable degradation of the physical properties of the polymer. Polythiophene, like polypyrrole, exhibits conductivity changes in response both to the amount of dopant and to the specific dopant, such as perchlorate, tetrafluoroborate, hexafluorophosphate, hydrogen sulfate, hexafluoroarsenate and trifluoromethylsulfonate.
Substituents placed on the heteroaromatic thiophene ring can affect the resulting conducting polymer. For example, thiophene polymerization can be affected by large substituents at the 3 and 4 positions, as seen in the inability of 3,4-dibromothiophene to polymerize. The electronic and steric effects introduced by the 3,4-dibromo substituents may prevent chain propagation. However, in contrast to pyrrole, ring substituents on thiophene do not seriously reduce the conductivity of the resulting heteroaromatic polymer. For example, it has been found that for 3-methylthiophene and 3,4-dimethylthiophene, the resulting substituted polythiophene exhibited an improved conductivity compared to the parent polythiophene, presumably due to enhanced order in the polymer chain of the substituted thiophene. However, the methyl group is not a suitable substituent for the subsequent polymer surface functionalization needed to produce an analyte sensor.
The following are representative references concerning the synthesis and conductivity of polythiophene and substituted polythiophenes:
From the studies on the polyacetylenes, polypyrroles and polythiophenes, and from related studies on other conducting polymers, including polyparaphenylene, polyazulene, polycarbazole, polypyrene, polyaniline and polytriphenylene, it is apparent that a delicate balance exists between the electronic effects and the steric effects introduced by the substituents that renders a polymer of a substituted five or six member heteroaromatic ring more conducting or less conducting than the unsubstituted parent heteroaromatic compound. Therefore, it would be advantageous to develop a monomer that can be readily polymerized, chemically or electrochemically, to yield a conducting polymer having sufficient conductivity such that the polymer can be used as an analyte sensor in a diagnostic device to determine the presence and concentration of an analyte in liquid media.
It is also apparent that a functionalized conducting polymer is required for ultimate use as an analyte sensor. The polymer must not only possess sufficient conductivity, but the polymer also must contain moieties that can interact with the analyte of interest. This interaction then must sufficiently alter the conductivity of the polymer in order to measurably detect the conductivity difference and convert this conductivity change into an analyte concentration. It is to such a conducting polymer that the method of the present invention is directed.
The prior art does not include any known references to the method of the present invention. The prior art chemical modifications to conducting polymers were unconcerned with the retention of high conductivity. For example, M. S. Wrighton et al, in the references cited above, have developed N-alkylpyrroles in an attempt to improve the binding of a polymer film to a platinum electrode. In this study, only a very thin layer of functionalized polypyrrole in contact with the electrode is required, therefore making the conductivity of the essential monolayer film unimportant.
Saloma et al (J. Electrochem. Soc. 132, 2379 (1985)) have attempted to functionalize polymer films in order to modify electrode properties. Saloma et al attempted to utilize the conductivity of the functionalized polymer as an electronic mediator for any chemical effects occurring on the attached moiety. However, this particular research area has been bypassed by similar chemical modifications of metal electrodes (R. W. Murray, Acc. Chem. Res. 13, 135 (1980)).
M. V. Rosenthal et al disclosed, in M. V. Rosenthal, T. A. Skotheim, C. Linkous and M. I. Florit, Polym. Preprints 25, 258 (1984) and in M. V. Rosenthal, T. A. Skotheim, J. Chem. Soc. Chem. Commun. 6, 342 (1985), an attempt to derivatize polypyrrole after polymerization.
The above referenced prior art concerning substituted pyrrole and substituted thiophene polymers is not directed to preparing conducting polymers for use as an analyte sensor in a diagnostic device. For example, films prepared from the methyl derivative of thiophene were not synthesized in order to attempt subsequent polymer surface functionalization, but rather to prevent monomeric couplings through the beta positions in order to introduce greater order, and therefore greater conductivity, into the polymer. In the referenced prior art, the investigators attempted to characterize and improve polymer properties, as opposed to chemically utilizing the substituents on the heteroaromatic ring.
During the course of the investigations on the synthesis and polymerization of functionalized 2,5-dithienylpyrrole derivatives, the electrochemical polymerization and the properties of the parent molecule, poly[2,5-di(2-thienyl)pyrrole], was disclosed by G. G. McLeod, M. G. B. Mahoubian-Jones, R. A. Pethuck, S. D. Watson, N. D. Truong, J. C. Galiri, and J. Francois in Polymer 27 (3), 455-8 (1986). The molecule, 2,5-di(2-thienyl)pyrrole (structure VI), is the parent heteroaromatic monomer that forms the basis of the method of the present invention. Although the primary objective of McLeod et al was to determine the solubility of the polymer resulting from 2,5-di(2-thienyl)pyrrole (VI), the polymerization of 2,5-di(2-thienyl)pyrrole was interesting for several additional reasons. For example, poly[2,5-di(2-thienyl)pyrrole] is readily synthesized electrochemically and, when anion doped, exhibits an electric conductivity analogous to polypyrrole and polythiophene. ##STR4##
However, most surprisingly and unexpectedly, and in accordance with the method of the present invention, 2,5-di(2-thienyl)pyrrole can be functionalized at the three-position of the pyrrole ring, and yield conducting polymers that exhibit the high conductivity of the unsubstituted parent dithienylpyrrole (VI). As will be discussed in the detailed description of the invention, a variety of functional groups can be incorporated into the three-position of the pyrrole ring of 2,5-di(2-thienyl)pyrrole without adversely affecting the conductivity of the resulting polymer.
In addition to the novel monomers used to synthesize the conducting polymers of the present invention, the conducting polymers can be further derivatized, after polymerization, to allow the detection and measurement of a specific analyte. According to the method of the present invention, postpolymerization derivatization and functionalization of the conducting polymer permits detection and measurement of a specific analyte by coupling the vibrational energy resulting from the functionalized polymer-analyte reaction to the phonon modes of the polymer. As used here, and throughout the specification, a phonon is a quantized, delocalized vibrational or elastic state of the polymer lattice.
Although several references disclose the use of conducting organic polymers in sensors, no known prior art references utilize the vibrational energy coupling of the analyte reaction to the conducting polymer. In fact, none of the present conducting polymer-based sensors involve an analyte probe molecule covalently bound to, and acting in concert with, the polymer. In contrast, the prior art sensors are based upon a direct interaction of an analyte, usually a gas, with the polymer. It should be noted however that the conducting polymers used in the present invention also can be used as an analyte sensor by direct interaction with the analyte.
The most common mode of direct interaction between the analyte and the conducting polymer is to affect the state of oxidation of the organic conducting polymer. As will be discussed more fully in the detailed description of the invention, the existence of bipolaron, and therefore the conductivity of the polymer, depends upon having the polymer oxidized, with the oxidation state supported by dopant counterions. Sensors then can be developed based upon either compensating conducting films or chemically doping reduced films.
For example, M. S. Wrighton et al, in European Pat. No. 185,941, discloses the use of conducting organic polymers as the active species in a chemical sensor. The patent generally teaches using the changes in physical properties of the conducting polymer as the active transduction into electrical signals. Specific examples cited in the patent include detection of oxygen gas, hydrogen gas, pH and enzyme substrate concentrations. The Wrighton et al patent neither teaches the coupling of an analyte/probe molecule vibrational interactions to the vibrational manifold of the polymer nor teaches the use of such vibrational coupling as a transduction mechanism for analyte detection. In contrast, the principal transduction mechanism described by Wrighton et al is the direct use of the change in polymer conductivity induced by oxidation or by reduction.
An additional mode of substrate/polymer interaction that is suitable for sensor development has been described in the prior art. It has been shown that it is possible to utilize the change of the surface dielectric attending the absorption of an analyte upon a polypyrrole film to make an alcohol sensor. In addition to novel electronic transduction mechanisms, the prior art also describes the use of a suspended gate, field effect transistor. Such electronic structures are in most ways analogous to well known structures employing inorganic semiconductors, and they can be expected to be generically useful in sensor development. In the embodiment of the invention described herein, a chemiresistor device configuration is used. It is anticipated, however, that evolutionary improvements will utilize the gated structures as described in the prior art.
The following references are representative of the state of the art of electrochemical sensors using heteroaromatic polymers:
Malmros, U.S. Pat. No. 4,444,892, disclosing a device having an analyte specific binding substance immobilized onto a semiconductive polymer to allow detection of a specific analyte.
European Pat. No. 193,154, filed Feb. 24, 1986, disclosing immunosensors comprising a polypyrrole or polythiophene film containing an occluded antigen or antibody.
M. Umana and J. Waller, Anal. Chem. 58, 2979 (1986) disclosed the occlusion, or trapping, of an enzyme, glucose oxidase, by electropolymerizing pyrrole in the presence of the enzyme. The polypyrrole containing the occluded enzyme then can be used to detect glucose. The method of the present invention however differs significantly in that according to the present invention the enzyme is covalently bound to the conducting polymer after polymerization.
The following references are cited to further show the state of the prior art and to serve as additional background material for the method of the present invention:
Vibrational energy transport in proteins:
A preferred synthesis of the parent molecule 2,5-di(2-thienyl)pyrrole:
The preparation of pyrrole derivatives 20 by 1,3-dipolar cycloaddition: